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Objectives: The Glucokinase Regulatory Protein GKRP, encoded by GCKR, enables acute regulation of liver glucokinase to support metabolic demand. The common human GCKR rs1260326:Pro446 > Leu variant within a large linkage disequilibrium region associates with pleiotropic traits including lower Type 2 diabetes risk and raised blood triglycerides and cholesterol. Whether the GCKR-P446 > L substitution is causal to the raised lipids is unknown. We determined whether mouse GKRP phenocopies the human GKRP:P446 > L substitution and studied a GKRP:P446L knockin mouse to identify physiological consequences to P446 > L. Methods: GKRP-deficient hepatocytes were transfected with adenoviral vectors for human or mouse GKRP:446 P or 446 L for cellular comprehensive analysis including transcriptomics consequent to P446 > L. Physiological traits in the diet-challenged P446L mouse were compared with pleiotropic associations at the human rs1260326 locus. Transcriptomics was compared in P446L mouse liver with hepatocytes overexpressing glucokinase or GKRP:446 P/L. Results: 1. P446 > L substitution in mouse or human GKRP similarly compromises protein expressivity of GKRP:446 L, nuclear sequestration of glucokinase and counter-regulation of gene expression. 2. The P446L knockin mouse has lower liver glucokinase and GKRP protein similar to human liver homozygous for rs1260326-446 L. 3. The diet-challenged P446L mouse has lower blood glucose, raised blood cholesterol and altered hepatic cholesterol homeostasis consistent with relative glucokinase-to-GKRP excess, but not raised blood triglycerides. Conclusions: Mouse GKRP phenocopies the human GKRP:P446 > L substitution despite the higher affinity for glucokinase of human GKRP. The diet-challenged P446L mouse replicates several traits found in association with the rs1260326 locus on chromosome 2 including raised blood cholesterol, lower blood glucose and lower liver glucokinase and GKRP protein but not raised blood triglycerides.

The Carbohydrate response element binding protein, ChREBP encoded by the MLXIPL gene, is a transcription factor that is expressed at high levels in the liver and has a prominent function during consumption of high-carbohydrate diets. ChREBP is activated by raised cellular levels of phosphate ester intermediates of glycolysis, gluconeogenesis and the pentose phosphate pathway. Its target genes include a wide range of enzymes and regulatory proteins, including G6pc, Gckr, Pklr, Prkaa1,2, and enzymes of lipogenesis. ChREBP activation cumulatively promotes increased disposal of phosphate ester intermediates to glucose, via glucose 6-phosphatase or to pyruvate via glycolysis with further metabolism by lipogenesis. Dietary fructose is metabolized in both the intestine and the liver and is more lipogenic than glucose. It also induces greater elevation in phosphate ester intermediates than glucose, and at high concentrations causes transient depletion of inorganic phosphate, compromised ATP homeostasis and degradation of adenine nucleotides to uric acid. ChREBP deficiency predisposes to fructose intolerance and compromised cellular phosphate ester and ATP homeostasis and thereby markedly aggravates the changes in metabolite levels caused by dietary fructose. The recent evidence that high fructose intake causes more severe hepatocyte damage in ChREBP-deficient models confirms the crucial protective role for ChREBP in maintaining intracellular phosphate homeostasis. The improved ATP homeostasis in hepatocytes isolated from mice after chronic activation of ChREBP with a glucokinase activator supports the role of ChREBP in the control of intracellular homeostasis. It is hypothesized that drugs that activate ChREBP confer a protective role in the liver particularly in compromised metabolic states.